The long-term goal of these studies is to understand the molecular mechanisms of herpesvirus nucleocapsid envelopment. All herpesviruses bud initially from the inner nuclear membranes of infected cells suggesting that information gained from these studies will be applicable to all members of this medically important virus family. The application focuses on the role of herpes simplex virus 1 UL31 and UL34 in the envelopment process. Hypotheses based on extensive preliminary data are tested. The first hypothesis proposes that the protein encoded by UL31 (pUL3 t) interacts with the UL34 gene product (pUL34) to mediate proper targeting of both proteins to the inner nuclear membrane. To assess the importance of the interaction in vivo, viral mutants bearing UL31 genes that lack pUL34 interacting domains, and UL34 mutants that lack pUL31 interacting regions will be tested for their abilities to correctly target both proteins in infected cells and to mediate nucleocapsid envelopment. The second hypothesis to be tested seeks to understand the role of nuclear lamina association of pUL31. The nuclear lamina is part of the nucleocytoskeleton or nuclear matrix that lines the inside surface of the nuclear rim. One possible role is that nuclear lamina association of pUL31 mediates anchoring of the pUL31/pUL34 complex at the nuclear rim. Therefore UL31 mutants lacking lamina association domains and containing known lamina binding domains replacing these domains will be tested for their abilities to mediate correct targeting of pUL31 and pUL34 to the nuclear rim in transient expression assays. Mutant viruses expressing these proteins will then be assessed for their ability to produce enveloped virions by electron microscopic examination of cells infected with these viruses. A second possible role is that pUL31 causes partial lamina depolymerization to allow nucleocapsids access to the inner nuclear membrane. To assess this possibility, the lamina of cells infected with wild type virus and a deletion virus lacking UL31 will be characterized by immunoelectron microsopy. If the wild type virus infected cells contain a porous lamina whereas cells infected with the UL31 mutant do not, mechanisms by which pUL31 could mediate local lamina depolymerization will be explored.